Recent studies have suggested that a minority of specialized cells in a number of adult tissues including brain and bone marrow are capable of assuming phenotype and function of cells from developmentally unrelated tissues. This observation has been termed plasticity and has potentially far-reaching implications for therapeutic applications. Controversy surrounds the question of whether the remarkable hematopoietic activity recovered from preparations of muscle tissue represents a similar example of developmental plasticity or merely indicates trafficking of marrow cells to muscle. To date this has only been examined in chimeric murine models generated using lethal irradiation. Resolution of this question is directly relevant to any potential application and modification of such cells. Therefore, the major objective of this proposal is to develop a sufficiently rigorous assay to determine the developmental origin of muscle-derived cells with hematopoietic potential without otherwise lethal irradiation of muscle donor tissue. We propose to use a unique murine model that relies on the selective and genetically correctable mitomycin C (MMC)-sensitivity of hematopoietic cells in a Fanconi Anemia mouse model. Thus, we will generate functionally chimeric nonlethally irradiated donor animals with a retrovirally corrected, MMC resistant and clonally marked hematopoietic compartment and a non-corrected muscle compartment for a series of transplantation studies. Importantly, our strategy combines a heritable, functionally distinct phenotype of muscle-derived versus marrow-derived hematopoietic progeny with retroviral marking for clonal tracking of engrafted cells. Independent of their developmental origin, the existence of hematopoietically active cells in muscle might conceivably allow the generation of autologous tissues for therapeutic applications with all the implicit advantages over conventional allogeneic grafts. We therefore additionally propose to study the phenotype and investigate the genetic modification of muscle-derived cells responsible for hematopoietic repopulation. Combined with recent insights into the molecular basis of a number of heritable diseases and substantial improvements in the design of vectors to genetically modify and phenotypically correct these cells, this may hold considerable therapeutic potential for the treatment of hereditary marrow failure syndromes including Fanconi Anemia, or as hematopoietic rescue in the context of high-dose therapy for malignancies.